Constructed wetland as a green remediation technology for the treatment of wastewater from underground coal gasification process.

The wastewater from underground coal gasification (UCG) process has extremely complex composition and high concentrations of toxic and refractory compounds including phenolics, aliphatic and aromatic hydrocarbons, ammonia, cyanides, hazardous metals and metalloids. So, the development of biological processes for treating UCG wastewater poses a serious challenge in the sustainable coal industry. The aim of the study was to develop an innovative and efficient wetland construction technology suitable for a treatment of UCG wastewater using available and low-cost media. During the bioremediation process the toxicity of the raw wastewater decreased significantly between 74%-99%. The toxicity units (TU) ranged from values corresponding to very high acute toxic for raw wastewater to non-toxic for effluents from wetland columns after 60 days of the experiment. The toxicity results correlated with the decrease of some organic and inorganic compounds such as phenols, aromatic hydrocarbons, cyanides, metals and ammonia observed during the bioremediation process. The removal percentage of organic compounds like BTEX, PAHs and phenol was around 99% just after 14 days of treatment. A similar removal rate was indicated for cyanide and metals (Zn, Cr, Cd and Pb). Concluded, in order to effectively assess remediation technologies, it is desirable to consider combination of physicochemical parameters with ecotoxicity measurements. The present findings show that wetland remediation technology can be used to clean-up the heavily contaminated waters from the UCG process. Wetland technology as a nature-based solution has the potential to turn coal gasification wastewater into usable recycled water. It is economically and environmentally alternative treatment method.

Hybrid nanolayers of star polymers and silver nanoparticles with antibacterial activity.

The aim of this study was to obtain stable star polymer layers with incorporated silver nanoparticles (AgNPs) and to study the antimicrobial activity of these hybrid materials. In this work, a novel approach regarding the synthesis of AgNPs directly by the star polymer layer is presented. Nanolayers of poly(N,N'-dimethylaminoethyl methacrylate) and hydroxyl-bearing poly[oligo(ethylene glycol) methacrylate] (P(DMAEMA-co-OEGMA-OH)) stars, covalently bound with solid supports, were obtained through chemical reaction of hydroxyl groups in the star arms with substrate modified with imidazole derivative. Quantitative chemical composition analysis and tracking of the changes in the morphology and wettability after every step of surface modification confirmed the covalent attachment of stars with the support. In the next step, the polymer nanolayers were modified with AgNPs formed in situ using only amine groups of the star arms and followed by the crystal quartz microbalance (QCM). The analysis of the layer thickness and affinity to water, both with the shape, size and amount of silver incorporated into the layer, confirmed the efficacy of AgNPs formation. The amount of silver incorporated into layers was correlated with the molar masses of the grafted stars, and a possible location of AgNPs within layers was shown. The antibacterial activity tests of prepared nanolayers showed that obtained hybrid materials were highly effective against both gram-positive and gram-negative bacteria strains. This study shows that the obtained layers are promising as stable coatings for antibacterial applications.

Insights into the microbial diversity and structure in a full-scale municipal wastewater treatment plant with particular regard to Archaea.

Due to limited description of the role and diversity of archaea in WWTPs, the aim of the study was to analyze microbial community structures and diversities with particular regard to Archaea in the samples taken from different stages of the full-scale municipal wastewater treatment plant and effluent receiving water (upstream and downstream discharge point). Our study was focused on showing how the treatment processes influenced the Eubacteria and Archaea composition. Alpha and Beta diversity were used to evaluate the microbial diversity changes in the collected samples. Proteobacteria was the largest fraction ranging from 28% to 67% with 56% relative abundance across all samples. Archaea were present in all stages of WWTP ranged from 1 to 8%. Among the Archaea, two groups of methanogens, acetoclastic (Methanosarcina, Methanosaeta) and hydrogenotrophic methanogens (Methanospirillium, Methanoculleus, Methanobrevibacter) were dominant in the technological stages. The obtained results indicate that the treated wastewater did not significantly affect eubacterial and archaeal composition in receiving water. However, differences in richness, diversity and microbial composition of Eubacteria and Archaea between the wastewater samples taken from the primary and secondary treatment were observed.

Seasonal and Technological Shifts of the WHO Priority Multi-Resistant Pathogens in Municipal Wastewater Treatment Plant and Its Receiving Surface Water: A Case Study.

The present study was focused on the identification of multi-resistant bacteria from the WHO priority pathogens list in the samples taken from different stages of the full-scale municipal wastewater treatment plant and receiving water. Additionally, the seasonal variations of the selected multi-resistant pathogens were analyzed in the samples. In order to the aim of the study, the metagenomic DNA from the collected samples was isolated and sequenced. The samples were collected in three campaigns (spring, summer, autumn). Metagenomic DNA was isolated by the commercial kits, according to the manufacturer's instruction. Illumina sequencing system was employed, and the R program was used to metagenomic analysis. It was found that the wastewater samples and receiving water contained the multi-resistant bacteria from the WHO priority pathogens list. The seasonal and technological variations affected the distribution of the pathogens in the wastewater. No effect of the effluent on the pathogens in the receiving water was observed. The results indicated that antibiotic-resistant "priority pathogens" from the WHO list are there in the waste- and receiving water. Technological process and seasons effected their distribution in the environment. Metagenomic analysis can be used as sufficient tool in microbiological and human health risk assessment.

Fluoroquinolone Resistance and Virulence Properties Among Wastewater Aeromonas caviae Isolates.

The study provides data on antibiotic resistance as well as the virulence characteristics of Aeromonas caviae isolated from raw and treated wastewater. The isolates were identified as A. caviae by 16S rRNA gene sequencing. In the analyzed strains, high frequency for the following genes was observed: aac(6')-Ib-cr, qnrB, and qnrD. The presence of qnrA and ogxB genes was not found in any strain. The higher frequency of the investigated genes was observed in strains from raw wastewater (RW). The strains of A. caviae showed multiple antibiotic resistance evaluated by the disk diffusion method. Multiple antibiotic resistance indices ranged from 0.36 to 0.69. Susceptibility to six heavy metals (Cd+2, Zn+2, Cu+2, Co+2, Mn+2, and Ni+2) was recorded for all the isolates. The order of metal resistance of A. caviae was Co > Cu > Zn > Cd > Ni > Mn. All the strains of A. caviae showed ß-hemolytic activity. Enzymes of amylase, cellulase, and lipase were produced by all isolates. Only the strains from RW had the ability to form biofilms and showed motility. The obtained results indicate that wastewater is a potential source and/or reservoir of virulent and multidrug-resistant A. caviae as "high-risk isolates."

A comprehensive study on bisphenol A degradation by newly isolated strains Acinetobacter sp. K1MN and Pseudomonas sp. BG12.

Bisphenol A (BPA) is an endocrine disrupting chemical. Its extensive use has led to the wide occurrence of BPA in various environmental ecosystems, at levels that may cause negative effects to the ecosystem and public health. Although there are many bacteria able to BPA utilization, only a few of them have a strong capacity for its biodegradation. Therefore, it is important to search for new bacteria strains, investigate their BPA biodegradation ability and potential effect of pH and other organic compounds on the process. These tasks have become the object of the present study. The results of our research show that for the newly isolated strains Acinetobacter sp. K1MN and Pseudomonas sp. BG12 after 15 days, with an initial BPA concentration of 100 mg L- 1, the highest BPA removal was achieved at pH 8, while sodium glutamate as a biostimulant best accelerated BPA degradation. Kinetic data for BPA biodegradation by both strains best fitted the Monod model. The specific degradation rate and the half saturation constant were estimated respectively as 8.75 mg L- 1 day- 1 and 111.27 mg L- 1 for Acinetobacter sp. K1MN, and 8.6 mg L- 1 day- 1 and 135.79 mg L- 1 for Pseudomonas sp. BG12. The half-maximal effective concentration (EC50) of BPA for Acinetobacter sp. K1MN was 120 mg L- 1 and for Pseudomonas sp. BG12 it was 123 mg L- 1. The toxicity bioassay (Microtox test) showed that elimination of BPA by both strains is accompanied by reduction of its toxic effect. The ability of tested strains to degrade BPA combined with their high resistance to this xenobiotic indicates that Acinetobacter sp. K1MN and Pseudomonas sp. BG12 are potential tools for BPA removal during wastewater treatment plant.

Whole-Genome Sequences of Antibiotic-Resistant Aeromonas caviae Strains Isolated from Treated Wastewater.

The presented data provide new information on antibiotic resistance and virulence genes in the genomes of Aeromonas caviae strains TW-2 and TW-6, isolated from treated wastewater. The results confirm the presence of multi-antibiotic-resistant Aeromonas caviae strains with virulence properties as "high-risk isolates" in treated wastewater.

Effect of the freeze-drying process on the phenotypic diversity of Pseudomonas putida strains isolated from the interior of healthy roots of Sida hermaphrodita: Phenotype microarrays (PMs).

The objective of this study was to research the effect of the freeze-drying process on the metabolic changes of Pseudomonas putida strains (E41, E42, R85) isolated from the interior of Sida hermaphrodita roots with the use of the phenotypic microarrays (PM) technology. The proposed method of the freeze-drying process with inulin as component lycoprotectant demonstrated a high bacterial survival ratio (BSR) immediately after freeze-drying and storage after 12 months. While, after 360 days of freeze-drying BSR decreased to value of 74.38. Pseudomonas putida strains were assayed on microplates PM1-PM5, and PM9-PM13 testing 664 different substrates. However, no significant differences in the use of C substrates were observed either before or after the freeze drying process. An insignificant negative effect of the freeze-drying on the use of these substrates was observed. The utilization of N, P and S sources was low or showed no metabolic activity for most of the compounds after freeze-drying. The freeze-drying process increased the sensitivity of the bacteria to antibiotics and selected chemicals. In this study, the freeze-drying process decreased the metabolic activities of the tested strains and their resistance to antibiotics and chemicals.

Properties of Antibiotic-Resistant Bacteria Isolated from Onsite Wastewater Treatment Plant in Relation to Biofilm Formation.

The aim of the present study was to determine some properties of antibiotic-resistant bacterial strains isolated from onsite wastewater technology in relation to biofilm formation, e.g., autoaggregation and motility. Additionally, biosurfactant production by the isolates was also evaluated. The ability of selected strains to develop a biofilm was assessed by using the crystal violet method, which allows to indirectly quantify the attached bacterial biomass (live, dead cells, and polysaccharides as well). Obtained results showed that 19 of the analyzed strains were able to produce biofilm after 72 h of incubation. The low values of surface tension in the range between 28 and 36 mN/m were observed in the bacteria, which are not able to produce biofilm or be classified as weak biofilm producers. Among biofilm-forming strains the highest autoaggregation index was observed for Mycobacterium brumae and Bacillus alcalophilus. Noteworthy, that some strains capable of biofilm formation showed no aggregation abilities or were characterized by low autoaggregative properties. The results of visual autoaggregation assay showed no visible flocs after given time of incubation. The results from motility test demonstrated that most of the analyzed strains were motile. Noteworthy, that up to now literature data about physiology, biofilm formation, and autoaggregative capabilities of bacteria isolated from onsite wastewater technology are very limited and this paper gives the information on the antibiotic-resistant bacteria with ability to form biofilm. Thus, the present study points to develop novel bioinocula in antibiotic degradation and to reach novel biofilm-dispersing agents produced by various bacteria that can be used as disinfectants or surface-coating agents to prevent microbial surface colonization and biofilm development.

Using phenotype microarrays in the assessment of the antibiotic susceptibility profile of bacteria isolated from wastewater in on-site treatment facilities.

The scope of the study was to apply Phenotype Biolog MicroArray (PM) technology to test the antibiotic sensitivity of the bacterial strains isolated from on-site wastewater treatment facilities. In the first step of the study, the percentage values of resistant bacteria from total heterotrophic bacteria growing on solid media supplemented with various antibiotics were determined. In the untreated wastewater, the average shares of kanamycin-, streptomycin-, and tetracycline-resistant bacteria were 53, 56, and 42%, respectively. Meanwhile, the shares of kanamycin-, streptomycin-, and tetracycline-resistant bacteria in the treated wastewater were 39, 33, and 29%, respectively. To evaluate the antibiotic susceptibility of the bacteria present in the wastewater, using the phenotype microarrays (PMs), the most common isolates from the treated wastewater were chosen: Serratia marcescens ss marcescens, Pseudomonas fluorescens, Stenotrophomonas maltophilia, Stenotrophomonas rhizophila, Microbacterium flavescens, Alcaligenes faecalis ss faecalis, Flavobacterium hydatis, Variovorax paradoxus, Acinetobacter johnsonii, and Aeromonas bestiarum. The strains were classified as multi-antibiotic-resistant bacteria. Most of them were resistant to more than 30 antibiotics from various chemical classes. Phenotype microarrays could be successfully used as an additional tool for evaluation of the multi-antibiotic resistance of environmental bacteria and in preliminary determination of the range of inhibition concentration.

Microbial Community Profiles in Wastewaters from Onsite Wastewater Treatment Systems Technology.

The aim of the study was to determine the potential of community-level physiological profiles (CLPPs) methodology as an assay for characterization of the metabolic diversity of wastewater samples and to link the metabolic diversity patterns to efficiency of select onsite biological wastewater facilities. Metabolic fingerprints obtained from the selected samples were used to understand functional diversity implied by the carbon substrate shifts. Three different biological facilities of onsite wastewater treatment were evaluated: fixed bed reactor (technology A), trickling filter/biofilter system (technology B), and aerated filter system (the fluidized bed reactor, technology C). High similarities of the microbial community functional structures were found among the samples from the three onsite wastewater treatment plants (WWTPs), as shown by the diversity indices. Principal components analysis (PCA) showed that the diversity and CLPPs of microbial communities depended on the working efficiency of the wastewater treatment technologies. This study provided an overall picture of microbial community functional structures of investigated samples in WWTPs and discerned the linkages between microbial communities and technologies of onsite WWTPs used. The results obtained confirmed that metabolic profiles could be used to monitor treatment processes as valuable biological indicators of onsite wastewater treatment technologies efficiency. This is the first step toward understanding relations of technology types with microbial community patterns in raw and treated wastewaters.

Application of the BIOLOG system for characterization of Serratia marcescens ss marcescens isolated from onsite wastewater technology (OSWT).

The scope of this study was to apply the Biolog system to identify and characterize a Serratia strain isolated from the surface of black plastic pieces which constitute the fluidized bed filter (onsite wastewater technology, OSWT). The preliminary isolation of the strain was done in the medium with tetracycline at a 16 mg/l concentration. To characterize the isolated strain, the following Biolog methods were applied: (1) EcoPlates microplates for evaluation of physiological profiling, (2) GEN III OmniLog® ID System for identification of the isolate, and (3) phenotypic microarrays (PM) technology for evaluation of sensitivity to antibiotics (PM11 and PM12). Results were recorded using the original OmniLog® software. The Serratia strain was identified as Serratia marcescens ss marcescens with similarity index 0.569. The same identification was obtained by the 16S rDNA analysis. PM analysis showed an enhancement of phenotype (resistance or growth) of this strain to 35 antibiotics. The loss of phenotype (sensitivity or non-growth) was observed only for 5 antibiotics: lomefloxacin (0.4 µg/ml), enoxacin (0.9 µg/ml), nalidixic acid (18.0 µg/ml), paromomycin (25.0 µg/ml) and novobiocin (1100 µg/ml). This study acknowledges that the methods proposed by the Biolog system allow correct and complete identification and characterization of the microbes isolated from different environments. Phenotypic microarrays could be successfully used as a new tool for identification of the multi-antibiotic resistance of bacteria and for determination of the minimal inhibition concentrations (MIC).